Crosslinking of interfacial layers in multilayered oil-in-water emulsions using laccase: Characterization and pH-stability

The enzymatic crosslinking of polymer layers adsorbed at the interface of oil-in-water emulsions was investigated. A sequential two step process, based on the electrostatic deposition of pectin onto a fish gelatin interfacial membrane was used to prepare emulsions containing oil droplets stabilized by fish gelatin-beet pectin membranes (citrate buffer, 10 mM, pH 3.5). First, a fine dispersed primary emulsion (5% soybean oil (w/v), 1% (w/w) gelatin solution) (citrate buffer, 10 mM, pH 3.5) was produced using a high pressure homogenizer. Second, a series of secondary emulsions were formed by diluting the primary emulsion into pectin solutions (0 - 0.4% (w/w)) to coat the droplets. Oil droplets of stable emulsions with different oil droplet concentrations (0.1%, 0.5%, 1.0% (w/v)) were subjected to enzymatic crosslinking. Laccase was added to the fish gelatin-beet pectin emulsions and emulsions were incubated for 15 min at room temperature. The pH- and storage stability of primary, secondary and secondary, laccase-treated emulsions was determined. Results indicated that crosslinking occurred exclusively in the layers and not between droplets, since no aggregates were formed. Droplet size increased from 350 to 400 nm regardless of oil droplet concentrations within a matter of minutes after addition of laccase suggesting formation of covalent bonds between pectin adsorbed at interfaces and pectin in the aqueous phase in the vicinity of droplets. During storage, size of enzymatically treated emulsions decreased, which was found to be due to enzymatic hydrolysis. Results suggest that biopolymer-crosslinking enzymes could be used to enhance stability of multilayered emulsions.